Pseudoknots
Overview While RNA is normally considered a linear nucleic acid structure, it can also have secondary RNA structures incorporated within the sequence. These structures are known as pseudoknots. Generally these structures stick up out of the linear sequence and come in a variety of shapes and sizes. The general ps eudoknot structure contains two helical regions that are connected together by single stranded regions or loops (1). The first pseudoknot structure was first identified in 1982 in the turnip yellow mosaic virus, a virus that infects insects (5). Pseudoknot Variations While there are a multitude of pseudoknot variations due to differences in the stem/loop length, there are 3 basic categories to define them by. Type B: Type B pseudoknots, otherwise known as bulge loop, involves base pairing between a single stranded region and a bulge (2). Type H: Type H pseudoknots, otherwise known as the hairpin loop, are the best characterized pseudoknots. They involve base-pairing between a single stranded region and the nucleotides in a hairpin loop (2). Type I: Type I pseudoknots, otherwise known as internal loop, involve interactions between a single stranded region and nucleotides within an internal loop (2). Functions and Examples Pseudoknots have a variety of functions in biological systems. Two of their most common roles include catalytical activity and inducing frameshift mutations. Catalytically Active Pseudoknots Pseudoknots play roles in biological activities such as telomerase function, removing introns, as well as forming the catalytic core of other various ribozymes in various organisms. A classic H-type pseudoknot is found within the core of human telomerase at the 5' end of the RNA sequence. This structure is required for the telomerase's primary function of maintaining chromosomal ends from degradation. Studies indicate that mutations found in the telomerase pseudoknot are directly correlated with various human diseases such as aplastic anemia and autosomal dyskeratosis congenita (1). In some eukaryotes, introns from the RNA sequence are self removed via self cleavage in order to form the correct translatable mRNA sequence. This group of introns are called group 1 self-splicing introns. A pseudoknot structure helps to establish the core of the group 1 self-splicing introns, by forming a "nest structure" that basepairs with all 3 domains of the ribozyme and therefore maintains its stability and catalytic function (1). In viral mRNA syntheis, particularly Hepatitis delta virus, the pseudoknot also aids in ribozyme catalytic activity. This virus has a circular genome that produces long strands of RNA sequences and therefore needs to splice it into smaller mRNA sequences to obtain individual proteins. This is done via self splicing at a ribozyme site that takes on the conformation of a double pseudoknot structure in order to be considered catalytically active and ready to cleave the RNA (1). Frameshift-Inducing Pseudoknots Framshift mutations are esstential for certain organisms to create altering quantities of proteins (ribosomal frameshift) or creating multiple proteins from the same piece of mRNA through open reading frames (polymerase frameshift). The efficiency of these frameshift inducing pseudoknots is also influenced by nearby sequences/structures as well as environmental elements (3). The main role of the pseudoknot in inducing polymerase based frameshifts is to cause the RNA polymerase that is reading the RNA to "slip" or stutter at the secondary structure's location and therefore reattach to the RNA at a different nucleotide in the sequence, thus leading to the frameshift. A similar system is seen in HIV-1 virus and is essential for its translational regulation. In this virus the pseudoknot is used to cause ribosomal frameshifting at a particular site between two genes within the mRNA sequence called the Gag and Pol junction. Over 95% of this junction has the ability to form 4 different pseudoknot variations. This causes the ribosome to slow down translation at this secondary structure (since its harder to translate physically) or cause the ribosome to dislodge and fall off the mRNA as well. This allows the virus to synthesis its Gag (structural) proteins and Pol (enzymatic) proteins at different ratios from eachother. For HIV-1, this is advantageous in the sense that the virus only produces the amount of each protein it needs, and doesn't waste time or energy accumulating uneeded proteins (4). This mechanism is seen in many other viruses as well as a way to regulate gene expression since they have to rely on host replication machinery. References 1. Staple, David W. "Pseudoknots: RNA Structures with Diverse Functions." PLOS Biology. PLOS Biology, 14 June 2011. 2. Burns, Robyn. "Pseudoknots." Nucleic Acids and Protein Synthesis. Oklahoma State University, 2006. 3. Gupta, A. "Local Structure and Enivronmental Factors Define the Effciency of an RNA Pseudoknot Involved in Programmed Ribosomal Frameshift Process." National Center for Biotechnology Information. U.S. National Library of Medicine, 3 Oct. 2014. PMID: 25226454 4. Huang, X. "Highly Conserved RNA Pseudoknots at the Gag-Pol Junction of HIV-1 Suggest a Novel Mechanism of -1 Ribosomal Frameshifting." National Center for Biotechnology Information. U.S. National Library of Medicine, May 2014. Pubmed ID: 24671765 5. "Pseudoknot" Wikepedia. Wikimedia